Blood processing apparatus with optical reference control

ABSTRACT

A density centrifuge blood processing system with automatic two-dimensional optical control of fluid separation by observing fluid characteristics in observation regions. The location of the regions is determined by monitoring an optical reference. Points representing edges of an optical reference are measured and lines are computed through the points. An error measurement is calculated for each line. If the error is too large, the image is abandoned. One of the lines is selected as a referent line. A new line is calculated orthogonal to the referent line. The error function is again computed for the dependant line. If the error exceeds a selected maximum, the frame is discarded. A transformation function translates data points from an (r, s) domain derived from measurements of the edges into an (x, y) domain used to identify pixels in the observation areas.

This patent application is a divisional of U.S. Ser. No. 12/233,185, nowU.S. Pat. No. 7,951,059, filed on Sep. 18, 2008, the contents of whichare incorporated herein by reference in their entirety.

FIELD OF THE INVENTION

The present invention relates to an apparatus and method for separatingparticles or components of a fluid. The invention has particularadvantages in connection with separating blood components, such asplatelets or white blood cells.

DESCRIPTION OF THE RELATED ART

In the medical field, it is often necessary to separate blood intocomponents. Whole blood consists of various liquid components andparticle components. The liquid portion of blood is largely made up ofplasma, and the particle components include red blood cells(erythrocytes), white blood cells (leukocytes), and platelets(thrombocytes). While these constituents have similar densities, theiraverage density relationship, in order of decreasing density, is asfollows: red blood cells, white blood cells, platelets, and plasma. Inaddition, the particle components are related according to size, inorder of decreasing size, as follows: white blood cells, red bloodcells, and platelets.

Typically, donated platelets are separated or harvested from other bloodcomponents using a centrifuge. White cells or other selected componentsmay also be harvested. The centrifuge rotates a blood separation vesselto separate components within the vessel or reservoir using centrifugalforce. In use, blood enters the separation vessel while it is rotatingrapidly and centrifugal force stratifies the blood components, so thatparticular components may be separately removed. Components are removedthrough ports arranged within stratified layers of blood components.

White blood cells and platelets in plasma form a medium-density,stratified layer or “buffy coat”. Because typical centrifuge collectionprocesses are unable to consistently and satisfactorily separate whiteblood cells from platelets in the buffy coat, other processes have beenadded to improve results. One separation process is one known ascentrifugal elutriation. In one common form of elutriation, a cell batchis introduced into a flow of liquid elutriation buffer, which carriesthe cell batch in suspension into a funnel-shaped chamber located on aspinning centrifuge. As additional liquid buffer solution flows throughthe chamber, the liquid sweeps smaller sized, slower-sedimenting cellstoward an elutriation boundary within the chamber, while larger,faster-sedimenting cells migrate to an area of the chamber having thegreatest centrifugal force.

When the centrifugal force and force generated by the fluid flow arebalanced, the fluid flow is increased to force slower-sedimenting cellsfrom an exit port in the chamber, while faster-sedimenting cells areretained in the chamber. If fluid flow through the chamber is increased,progressively larger, faster-sedimenting cells may be removed from thechamber.

The apparatus has a fluid separation chamber having a firstfrustro-conical segment adjacent a fluid inlet and radially inwardtherefrom, a second frustro-conical segment immediately adjacent thefirst frustro-conical segment and radially inward therefrom, the secondfrustro conical segment having a taper such that particles within thesecond frustro-conical segment are subjected to substantially equal andopposite centripetal and fluid flow forces. The taper of the secondfrustro-conical segment is selected based on the expected size ofparticles, such that at least particles of the average size of expectedparticles will be subjected to substantially equal and oppositecentripetal and fluid forces. The apparatus has at least one pumpcontrolling a rate of fluid flow through the fluid separation chamber, acamera configured to observe fluid flow with respect to the fluidseparation chamber, and a controller receiving signals from the cameraand controlling the motor and the pump.

For these and other reasons, there is a need to improve control ofparticle separation and/or separation of components of a fluid.

Additional technology related to this application is disclosed in, forexample, U.S. Pat. No. 5,722,926, issued Mar. 3, 1998; U.S. Pat. No.5,951,877, issued Sep. 14, 1999; U.S. Pat. No. 6,053,856, issued Apr.25, 2000; U.S. Pat. No. 6,334,842, issued Jan. 1, 2002; U.S. patentapplication Ser. No. 10/905,353, filed Dec. 29, 2004; U.S. patentapplication Ser. No. 11/163,969, filed Nov. 4, 2005 and in particularU.S. Pat. No. 7,422,693, filed Jul. 1, 2004.

SUMMARY OF THE INVENTION

The present invention comprises a blood component separation apparatushaving a rotor for centrifugally separating blood into phases such asred blood cells, buffy coat, or plasma. A camera monitors a separationchamber and image processing determines the location of boundaries. Theapparatus controls the position of the boundaries by adjusting the speedof pumps or the rotor or both.

In the present invention, fluid flow in a blood separation chamber in acentrifugal separation device is selectively controlled by opticalsensing of two regions in the separation chamber. Interface position maybe controlled by optical sensing of a two-dimensional view of theinterface in the separation chamber in an area adjacent an outflow portor ports. Gross adjustments, that is, relatively large changes in thelocation of the interface or interfaces are best controlled by thisobservation of the interface. Thus in transient states, such as theinitial setup of flow conditions, interface position sensing can beeffective. Fluid flow may also be controlled in response to the opticalintensity (light or dark) of the fluid in the outflow tube. This opticalintensity correlates to presence of certain blood components such as redblood cells. Fine adjustments, that is, relatively small changes in thelocation of the interface are best controlled by sensing the opticalintensity in the outflow tube. Thus in steady state conditions, such asthe extraction of a blood component through the outflow tube, outflowintensity sensing is more effective.

In a high-speed centrifuge for separating blood components, imaging ofthe same locations for observation regions from rotation to rotationpresents significant problems, particularly in view of the vibrationswith high-speed rotation. The present apparatus controls the interfacelocation by measuring light intensity in the collect port monitoringregion in the collect port by detecting the presence or absence of RBC'sin the collect port, and by monitoring the interface in the phaseboundary or interface monitoring region. In order for the apparatus tocontrol the interface a reference position on the disposable bloodprocessing bag, which is carried on the rotor, must be rapidly andreliably determined. In this invention, this is accomplished by adetection algorithm, which monitors an L-shaped calibration marker oroptical reference. An intersection derived from edges of the opticalreference is used as an origin. A series of points representing an edgeis measured. A set of data points, preferably about five (5), iscollected for each edge, and a line is computed through the points. Anerror measurement is calculated for each line. If the error is toolarge, the image (an “observation” or “frame”) for the current rotationis abandoned. The line with the least error is selected as a referentline. A new or dependant line is calculated for the line with thegreater error. The error function is again computed for the dependantline. If the error exceeds a selected maximum, the observation or frameis discarded.

Using the parameters of the lines a transformation function is produced,which translates data points from an observation from an (r, s)co-ordinate domain derived from measurements of the edges into an (x, y)co-ordinate domain used to identify pixels in the observation areas. Totest the transformation, the data points for the two edges aretranslated from the (r, s) domain into the (x, y) domain and the errorfunction is computed once again. If the error exceeds the maximum errorlimit, the frame is abandoned.

If the data passes the tests, the pixels falling within the observationregions identified with reference to the origin that has been identifiedas the intersection of the referent and dependant lines are useddetermine the position of phase boundaries and out flow characteristics.The process outlined above and described more completely hereafterallows for a frame by frame determination of the location of an originin the pixel field of the camera and for a determination that the imageis sufficiently clear for the collection of data. Vibration and relativemotion between the rotor and separation chamber and the camera causesthe image detected by the camera to move in the (x, y) plane and to comein and out of focus. The method described allows the apparatus todiscard a frame or observation that is too blurry to provide accuratedata and to locate a consistent origin from frame to frame.

It is an object of the present invention to provide a density centrifugeblood processing system for separating fluid components comprising aseparation chamber rotating about a central rotation axis, theseparation chamber having an optical reference mounted thereon, and acomputational apparatus distinguishing the optical reference andestablishing reference coordinates for gathering data from at least oneobservation of the observation region detected by the first detector.

It is also an object of the invention that computational apparatusrejects an observation of the separation chamber if an error measurementof the optical reference is not within a pre-selected error.

A further object of the invention is to translate data from theobservation of the observation region from a first co-ordinate domaininto a second co-ordinate domain.

Another object is to provide an optical reference comprising at leasttwo non-parallel sides and wherein said computational apparatusrecognizes a first side represented by a first line and further fits asecond line to a second side according to a known angle between saidfirst side and said second side and to reject an observation of theseparation chamber if data representing an edge of the optical referenceare not within a pre-selected error.

It is also an object of the invention to compute an error measurementfor each of the first and second sides and to select the side with theleast error as a referent line, to compute a dependant line for the sidewith the greater error, to re-compute an error measurement for thedependant line, and to reject an observation of said separation chamberif the error measurement for the dependant line is not within apre-selected error.

These and other objects and features of the present invention will beapparent from the following detailed description. It is to be understoodthat both the foregoing general description and the following detaileddescription are exemplary, and are intended to provide furtherexplanation of the invention as claimed.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a partial perspective, schematic view of a blood processingcentrifuge apparatus including a fluid chamber.

FIG. 2 is a partial perspective, schematic view of the centrifugeapparatus and a control camera.

FIG. 3 is a partial cross-sectional view of blood processing apparatusof FIG. 2, including the fluid chamber of FIG. 1.

FIG. 4 is a partial cross-sectional, schematic view of a portion of aseparation vessel and the fluid chamber mounted on a centrifuge rotor ofFIG. 1.

FIG. 5 is a plan view of a separation chamber of the separation vesselof FIG. 4.

FIG. 6 is a graphic representation of steps for image processingaccording to the present invention.

FIG. 7 is a diagram showing the relationship of an (r, s) co-ordinatesystem and an (x, y) co-ordinate system.

FIG. 8 is a graph of signals for recognizing data points representing anedge of an optical reference.

DETAILED DESCRIPTION

The present invention preferably comprises a blood processing apparatushaving a camera control system, as disclosed in U.S. Pat. No. 7,422,693and in U.S. application Ser. Nos. 10/905,353 and 11/772,692 and11/774,073. It may also be practiced with a TRIMA® blood componentcentrifuge manufactured by CaridianBCT, Inc. of Colorado (formerlyGambro BCT, Inc.) or, alternatively, with a COBE® SPECTRA single-stageblood component centrifuge also manufactured by CaridianBCT, Inc. Boththe TRIMA® and the SPECTRA centrifuges incorporate a one-omega/two-omegasealless tubing connection as disclosed in U.S. Pat. No. 4,425,112 toIto. The SPECTRA centrifuge also uses a single-stage blood componentseparation channel substantially as disclosed in U.S. Pat. No. 4,094,461to Kellogg et al. and U.S. Pat. No. 4,647,279 to Mulzet et al. Theinvention could also be practiced with a TRIMA® or TRIMA ACCEL®centrifugal separation system or other types of centrifugal separator.The method of the invention is described in connection with theaforementioned blood processing apparatus and camera control system forpurposes of discussion only, and this is not intended to limit theinvention in any sense.

As embodied herein and illustrated in FIG. 1, a centrifuge apparatus 10has a centrifuge rotor 12 coupled to a motor 14 so that the centrifugerotor 12 rotates about its axis of rotation A-A. The motor 14 is coupledto the rotor 12 directly or indirectly through a shaft 24 connected tothe rotor 12. Alternately, the shaft 24 may be coupled to the motor 14through a gearing transmission (not shown).

The rotor 12 has a retainer 16 including a passageway or annular groove18 having an open upper surface adapted to receive a separation vessel28, shown in pertinent part in FIG. 4. The groove 18 completelysurrounds the rotor's axis of rotation A-A and is bounded by an innerwall 20 and an outer wall 22 spaced apart from one another to define thegroove 18. Although the groove 18 shown in FIG. 1 completely surroundsthe axis of rotation A-A, the groove could partially surround the axisA-A if the separation vessel is not annular. Preferably, a substantialportion of the groove 18 has a constant radius of curvature about theaxis of rotation A-A and is positioned at a maximum possible radialdistance on the rotor 12. This shape ensures that substances separatedin the separation vessel 28 undergo relatively constant centrifugalforces as they pass from an inlet portion to an outlet portion of theseparation vessel 28.

As shown in FIG. 1, a bracket 26 is provided on a top surface of therotor 12. The bracket 26 releasably holds a fluid chamber 30 on therotor 12 so that an outlet 32 of the fluid chamber 30 is positionedcloser to the axis of rotation A-A than an inlet 34 of the fluid chamber30. The bracket 26 preferably orients the fluid chamber 30 on the rotor12 with a longitudinal axis of the fluid chamber 30 in a planetransverse to the rotor's axis of rotation A-A. In addition, the bracket26 is preferably arranged to hold the fluid chamber 30 on the rotor 12with the fluid chamber outlet 32 facing the axis of rotation A-A.Although the fluid chamber 30 is shown on a top surface of the rotor 12,the fluid chamber 30 could also be secured to the rotor 12 at alternatelocations, such as beneath the top surface of the rotor 12.

FIG. 2 schematically illustrates an exemplary embodiment of an opticalmonitoring system 40 capable of measuring a distribution of scatteredand/or transmitted light intensities corresponding to patterns of lightoriginating from an observation region on the separation vessel 28. Themonitoring system 40 comprises light source 42, light collection element44, and detector 46. Light source 42 is in optical communication withthe centrifuge apparatus 10 comprising rotor 12, which rotates aboutcentral rotation axis A-A. Rotation about central rotation axis A-Aresults in separation of a blood sample in the separation vessel 28 intodiscrete blood components.

Light source 42 provides incident light beam 54, which stroboscopicallyilluminates an observation region 58 when the observation region 58passes under the light collection element 44. Light source 42 is capableof generating an incident light beam, a portion of which is transmittedthrough at least one blood component undergoing separation in separationvessel 28. At least a portion of scattered and/or transmitted light 56from the observation region 58 is collected by light collection element44. Light collection element 44 is capable of directing at least aportion of the collected light 56 onto detector 46. The detector 46detects patterns of scattered and/or transmitted light 56 from theobservation region. The observation region 58 may also be illuminated byan upper light source 62, which is positioned on the same side of theseparation chamber as the light collection element 44 and detector 46.Upper light source 62 is positioned such that it generates an incidentbeam 64, which is scattered by the blood sample and/or centrifuge. Aportion of the light from upper light source 62 is collected by lightcollection element 44 and detected by detector 46, thereby measuring adistribution of scattered and/or transmitted light intensities.

Distributions of scattered and/or transmitted light intensities compriseimages corresponding to patterns of light originating from theobservation region 58. The images may be monochrome images, whichprovide a measurement of the brightness of separated blood componentsalong the separation axis. Alternatively, the images may be colorimages, which provide a measurement of the colors of separated bloodcomponents along the separation axis. Observation region 58 ispositioned on a portion of the density centrifuge 10, preferably on theseparation vessel 28. The fluid chamber 30 may also be an observationregion, as explained below. In the exemplary embodiment illustrated inFIG. 4 and FIG. 5, separated blood components and phase boundariesbetween optically differentiable blood components are viewable inobservation region 58.

Detector 46 is also capable of generating output signals correspondingto the measured distributions of scattered and/or transmitted lightintensities and/or images. The detector 46 is operationally connected toa device controller 60 capable of receiving the output signals. Devicecontroller 60 displays the measured intensity distributions, stores themeasured intensity distributions, processes measured intensitydistributions in real time, transmits control signals to various opticaland mechanical components of the monitoring system and centrifuge or anycombination of these. Device controller 60 is operationally connected tocentrifuge apparatus 10 and is capable of adjusting selected operatingconditions of the centrifuge apparatus, such as the flow rates ofcellular and non-cellular components out of the separation vessel 28 orfluid chamber 30, the position of one or more phase boundaries,rotational velocity of the rotor about central rotation axis A-A, theinfusion of anticoagulation agents or other blood processing agents tothe blood sample, or any combination of these.

Device controller 60 can also be operationally connected to light source42 and/or upper light source 62. Device controller 60 and/or detector 46are capable of generating output signals for controlling illuminationconditions. For example, output signals from the detector 46 can be usedto control the timing of illumination pulses, illumination intensities,the distribution of illumination wavelengths and/or position of lightsource 42 and/or upper light source 62. Device controller 60 anddetector 46 are in two-way communication, and the device controllersends control signals to detector 46 to selectively adjust detectorexposure time, detector gain and to switch between monochrome and colorimaging.

Light sources comprise light emitting diode (LED) sources capable ofgenerating one or more incident beams for illuminating an observationregion on the centrifuge. A plurality of lamps may be positioned toilluminate a single side or multiple sides of the centrifuge apparatus10. Light emitting diodes and arrays of light emitting diode lightsources are preferred for some applications because they are capable ofgenerating precisely timed illumination pulses. Preferred light sourcesgenerate an incident light beam having a substantially uniformintensity, and a selected wavelength range.

The optical monitoring system comprises a plurality of light sources,each capable of generating an incident light beam having a differentwavelength range, for example, a combination of any of the following:white light source, red light source, green light source, blue lightsource and infra red light source. Use of a combination of light sourceshaving different wavelength ranges is beneficial for discriminating andcharacterizing separated blood fractions because absorption constantsand scattering coefficients of cellular and non-cellular components ofblood vary with wavelength. For example, a component containing redblood cells is easily distinguished from platelet-enriched plasma byillumination with light having wavelengths selected over the range ofabout 500 nm to about 600 nm, because the red blood cell componentabsorbs light over this wavelength significantly more strongly that theplatelet-enriched plasma component. In addition, use of multiple coloredlight sources provides a means of characterizing the white blood celltype in an extracted blood component. As different white blood celltypes have different absorption and scattering cross sections atdifferent wavelengths, monitoring transmitted and/or scattered lightfrom a white cell-containing blood component provides a means ofdistinguishing the various white blood cell types in a blood componentand quantifying the abundance of each cell-type.

The light sources provide a continuous incident light beam or a pulsedincident light beam. Pulsed light sources are switched on and offsynchronously with the rotation of the rotor 12 to illuminate anobservation region having a substantially fixed position on the rotor12. Alternatively, pulsed light sources of the present invention can beconfigured such that they can be switched on and off at differentangular positions, synchronous with the rotation of the rotor 12,illuminating different observation regions for each full rotation. Thisalternative embodiment provides a method of selectively adjusting thelocation of the observation region and, thereby, probing differentregions of the separation chamber 28 or of the fluid chamber 30.Triggering of illumination pulses may be based on the rotational speedof the centrifuge or on the angular position of the separation chamberor the fluid chamber 30 as detected by optical or electronic methodswell known in the art. Triggering may be provided by trigger pulsesgenerated by the device controller 60 and/or detector 46.

FIG. 3 is a cutaway view of the optical monitoring system 40. Theillustrated optical monitoring system 40 comprises CCD (“charge-coupleddevice”) camera 72 (CMOS (“complementary metal oxide semiconductor”) orother cameras could also be used) equipped with a fixed focus lenssystem (corresponding to the light collection element 44 and detector46), an optical cell 74 (corresponding to the observation region 58), anupper LED light source 76 (corresponding to the upper light source 62),and a bottom pulsed LED light source 78 (corresponding to the lightsource 42). As illustrated in FIG. 3, CCD camera 72 is in opticalcommunication with optical cell 74 and positioned to intersect opticalaxis 80. Upper LED light source 76 is in optical communication withoptical cell 74 and is positioned such that it is capable of directing aplurality of collimated upper light beams 82, propagating alongpropagation axes that intersect optical axis 80, onto the top side 84 ofoptical cell 74. Bottom pulsed LED light source 78 is also in opticalcommunication with optical cell 74 and is positioned such that it iscapable of directing a plurality of collimated bottom light beams 86,propagating along optical axis 80, onto the bottom side 88 of opticalcell 74.

CCD camera 72 may be positioned such that the focal plane of the fixedfocus lens system is substantially co-planar with selected opticalsurfaces of optical cell 74, such as optical surfaces corresponding toan interface monitoring region, calibration markers, one or moreextraction ports and one or more inlets. The CCD camera 72 is separatedfrom the center of the fixed focus lens system by a distance alongoptical axis 80 such that an image corresponding to selected opticalsurfaces of optical cell 74 is provided on the sensing surface of theCCD camera. This optical configuration allows distributions of lightintensities comprising images of rotating optical cell 74 or of fluidchamber 30 to be measured and analyzed in real time.

Referring to FIG. 3, first transparent plate 96 is provided between CCDcamera 72 and optical cell 74, and second transparent plate 98 isprovided between bottom LED light source 78 and optical cell 74. Firstand second transparent plates 96 and 98 physically isolate CCD camera72, upper LED light source 76 and bottom LED light source 78 fromoptical cell 74 so that these components will not contact a sampleundergoing processing in the event of sample leakage from the separationchamber 28. In addition, first and second transparent plates 96 and 98minimize degradation of CCD camera 72, upper LED light source 76 andbottom LED light source 78 due to unwanted deposition of dust and othercontaminants that can be introduced to the system upon rotation of theseparation chamber and filler. Further, first and second transparentplates 96 and 98 also allow a user to optimize the alignment of thecamera 72, upper LED light source 76 and bottom LED light source 78without exposure to a blood sample in the separation chamber 28. Firstand second transparent plates 96 and 98 can comprise any materialcapable of transmitting at least a portion of upper and bottomillumination light beams 82 and 86. Exemplary materials for first andsecond transparent plates 96 and 98 include, but are not limited to,glasses such as optical-quality, scratch-resistant glass, transparentpolymeric materials such as transparent plastics, quartz, or inorganicsalts.

FIG. 4 schematically illustrates the separation vessel 28 and fluidchamber 30 mounted on the rotor 12. The separation vessel 28 has agenerally annular flow path 100 and includes an inlet portion 102 andoutlet portion 104.

A radial outer wall 108 of the separation vessel 28 is positioned closerto the axis of rotation in the inlet portion 102 than in the outletportion 104. During separation of blood components, this arrangementcauses formation of a very thin and rapidly advancing red blood cell bedin the separation vessel 28 between the inlet portion 102 and outletportion 104. The red blood cell bed substantially limits or preventsplatelets from contacting the radial outer wall 108 of the separationvessel 28. This is believed to reduce clumping of platelets caused whenplatelets contact structural components of centrifugal separationdevices.

The inlet portion 102 includes an inflow tube 110 for conveying a fluidto be separated, such as whole blood, into the separation vessel 28.During a separation procedure, substances entering the inlet portion 102follow the flow path 100 and stratify according to differences indensity in response to rotation of the rotor 12. The outlet portion 104includes first, second, and third outlet lines 112, 114, 116 forremoving separated substances from the separation vessel 28. Preferably,each of the components separated in the vessel 28 is collected andremoved in only one area of the vessel 28, namely the outlet portion104. In addition, the separation vessel 28 preferably includes asubstantially constant radius except in the region of the outlet portion104 where the outer wall of the outlet portion 104 is preferablypositioned farther away from the axis of rotation to allow for outletports of the lines 112, 114, and 116 to be positioned at differentradial distances and to create a collection pool with greater depth forthe high density red blood cells. The outlet port of line 114 is fartherfrom the axis of rotation A-A than the other ports to remove higherdensity components, such as red blood cells. The port of line 116 islocated closer to the axis of rotation than the other ports to removethe least dense components separated in the separation vessel 28, suchas plasma. The first line 112 collects intermediate density componentsand, optionally, some of the lower density components. The first line112 may be coupled to the inlet 34 of the elutriation chamber 30. Theoutlet 32 of the elutriation chamber 30 is coupled to a line 130. Thesecond and third lines 114 and 116 are positioned downstream (not shown)from first line 112 to collect the high and low density components.

The positions of the interfaces are controlled by the CCD camera 72monitoring the position of the interface and controlling flow of liquidand/or particles in response to the monitored position. Further detailsconcerning the structure and operation of the separation vessel 28 aredescribed in U.S. Pat. No. 7,422,693 and also in U.S. Pat. No. 4,094,461to Kellogg et al. and U.S. Pat. No. 4,647,279 to Mulzet et al.

Referring to FIG. 2, the camera 44 is generally focused on theseparation vessel 28 and stroboscopic illumination allows an observationregion 58 around the first, second, and third lines 112, 114, and 116 tobe observed. Using information gathered through the camera, thecontroller 60 regulates the position of interfaces between various bloodcomponents, such as plasma, buffy coat (containing monocytes and/orwhite blood cells and platelets) and red blood cells by controlling thepumps (not shown) connected to lines 110, 114, 116, and 130. FIG. 5shows an image of the observation region 58 generated by the methods ofU.S. Pat. No. 7,422,693 corresponding to the separation of a human bloodsample and extraction of a separated platelet-containing bloodcomponent. The observation region 58 shown in FIG. 5 includes a phaseboundary monitoring region 202 and an extraction or collect portmonitoring region 204. Visible in phase boundary monitoring region 202are a red blood cell component 206, a plasma component 208 and amixed-phase buffy coat layer 210, which has both white blood cells andplatelets. Calibration markers are also apparent in the image in FIG. 5.Near an edge 212 of the optical cell 74 is an L-shaped calibrationmarker or optical reference 214 for determining the absolute position ofphase boundaries between optically differentiable blood components.Inner edges 232, 234 of the optical reference 214 are used to indicatethe positions and physical dimensions of the phase boundary monitoringregion 202 and the platelet collect port monitoring region 204. Thephysical dimension may be determined by adjusting the optics to within aselected range and then configuring the software with a parameter toconvert pixels to microns. Alternatively, the thickness of the opticalreference, usually about 1 mm, could be used. Light intensitiestransmitted through the phase boundary monitoring region 202 areacquired as a function of time and analyzed in real time to providemeasurements of the position of the phase boundary 216 between red bloodcell component 206 and buffy coat layer 210 and the phase boundary 218between the buffy coat layer 210 and plasma component 208. All boundarylayer positions are measured relative to the edge of the opticalreference 214.

Collect port monitoring region 204 monitors flow in first line 112 ofthe optical cell for extracting a blood component, for example, buffycoat. The apparatus responds to changes in detected blood component flowto establish a correct phase boundary level and further responds tochanges in observed phase boundaries to maintain a consistent phaseboundary level. The system discriminates between a plasma flowcondition, a buffy coat flow condition, and a red blood cell flowcondition, and can detect pump-induced flow variation in the bloodcomponent flow in the collect port measuring area. A plasma signal limitand a red blood cell signal limit may be set and the flow of fluidadjusted based on said limits. The system derives a statistical measureof fluid flow in the collect port measuring area, which may be a movingmedian of the average value of intensity of pixels in the collect portmeasuring area.

In this example, first line 112 having orifice 224 is configured tocollect buffy coat in the human blood sample and extends a distancealong the separation axis such that it terminates proximate to the buffycoat layer in the rotating separation chamber. The two-dimensionaldistribution of light intensities of light transmitted through thecollect port in the collect port monitoring region 204 depends on theconcentration, and spatial distribution and cell-type of cellularmaterial exiting the separation chamber. Light intensities transmittedthrough the collect port monitoring region 204 are acquired as afunction of time and analyzed to characterize the composition and fluxof cellular material out of the separation chamber 28. As cellularmaterials, such as white blood cells and red blood cells, absorb andscatter light from the light sources, passage of cellular materialthrough the extraction port decreases the observed light intensities.

The first collection line 112 is connected to the fluid chamber inlet 34to pass the intermediate density components into the fluid chamber 30.Components initially separated in the separation vessel 28 are furtherseparated in the fluid chamber 30. For example, white blood cells couldbe separated from plasma and platelets in the fluid chamber 30. Thisfurther separation preferably takes place by forming a saturatedfluidized bed of particles in the fluid chamber 30. The fluid chamber 30may be formed of a transparent or translucent co-polyester plastic, suchas PETG, to allow viewing of the contents within the chamber interiorwith the aid of the camera during a separation procedure.

The apparatus 10 includes the controller 60 (FIG. 1) connected to themotor 14 to control rotational speed of the rotor 12. The controller 60is connected to the pumps in lines 110, 114, 116, and 130 to control theflow rate of substances flowing to and from the separation vessel 28 andthe fluid chamber 30. The controller 60 controls the operation and flowrate of the pumps to permit the temporary purging of the fluid chamber30. The controller 60 may include a computer having programmedinstructions provided by a ROM or RAM as is commonly known in the art.The controller 60 may vary the rotational speed of the centrifuge rotor12 by regulating frequency, current, or voltage of the electricityapplied to the motor 14. Alternatively, the rotational speed can bevaried by shifting the arrangement of a transmission (not shown), suchas by changing gearing to alter a rotational coupling between the motor14 and rotor 12. The controller 60 may receive input from a rotationalspeed detector (not shown) to constantly monitor the rotation speed ofthe rotor.

Accumulated buffy coat components, comprising platelets, some whiteblood cells, and plasma, are removed via the first collection line 112.As the platelets, plasma, white blood cells, and possibly a small numberof red blood cells pass through the first collection line 112, thesecomponents flow into the fluid chamber 30, filled with the primingfluid, so that a saturated fluidized particle bed may be formed. Theplatelets flow toward the first collection line 112. The priming fluidalong the inner walls of the separation vessel 28 reduces the effectivepassageway volume and area in the separation vessel 28 and therebydecreases the amount of blood initially required to prime the system ina separation process. The reduced volume and area also induces higherplasma and platelet velocities next to the stratified layer of red bloodcells, in particular, to “scrub” platelets toward the first collectionline 112. The rapid conveyance of platelets increases the efficiency ofcollection.

The fluid chamber 30 is configured to allow cyclic collection ofselected particles, such as platelets, followed by efficient evacuationof the cells into a collection bag. In contrast to other chamber designsfor forming saturated fluidized beds, the fluid chamber described hereinhas particular application for the automated collection of bloodcomponents in that a bolus of cells having selected characteristics canbe collected in the fluid chamber 30 and then flushed with low densityfluid into a collection bag and these steps can be repeated multipletimes, allowing a larger quantity of the selected cells to be collectedfrom the donor or patient while reducing the amount of time necessaryfor the donation process. Collection of cells in the fluid chamber canbe monitored by the camera 72 and the device controller 60. When aselected quantity of cells have been collected in the fluid chamber 30,the flow of plasma through the chamber can be increased and thecollected cells can be washed out of the chamber and directed into acollection bag.

Frame-by-Frame Origin Calibration

In a high-speed centrifuge for separating blood components, control ofthe interface between blood components presents significant controlproblems. The present apparatus controls the interface location bymeasuring light intensity in the collect port monitoring region 204 inthe collect port by detecting the presence or absence of RBC's in thecollect port, and by monitoring the interface 216 or 218 in the phaseboundary or interface monitoring region 202. The light intensity in thecollect port can be measured by both an average value over a relativelybrief period of time or by a median value over a longer period of timeor by a combination of both measurements. The location of the interfaceis detected by a series of image processing steps, which allow theapparatus to recognize a boundary or interface despite limitations suchas the high speed of the centrifuge rotor, the characteristics ofstroboscopic light used for observation, or the limits of dataprocessing time. Monitoring the interface in the interface monitoringregion 202 allows the apparatus to determine and control the location ofthe interface reliably. In order for the apparatus to control theinterface, a reference position on the disposable blood processing bag,which is carried on the rotor, must be rapidly and reliably determined.In this invention, this is accomplished by a detection algorithm 230,which monitors the L-shaped calibration marker or optical reference 214.The detection algorithm 230 is shown in FIGS. 6 a through 6 f.

An optically controlled centrifuge for blood separation, as describedherein, presents certain problems for the control of the apparatus. Witha camera mounted on the frame and observing indistinct phenomena on arotor spinning in excess of 3000 rpm, vibration is a persistent problem.As viewed through the camera, the image of the separation chambershakes. Moreover, distances between the camera and observation areascannot be controlled to the requisite tolerances for observationpurposes. This is particularly true where, as herein, the blood is to beprocessed in a disposable blood separation chamber. Consequently, thelocation of observation regions must be determined dynamically. In thepresent invention, an optical reference having at least two non-paralleledges is mounted near the separation chamber. An intersection derivedfrom edges is determined for use as an origin. Observation regions areestablished with respect to the origin and pixels falling within theobservation regions are used to detect phase boundaries and outflowconditions for controlling the blood processing apparatus, as more fullydescribed in U.S. application Ser. No. 11/772,692 and U.S. applicationSer. No. 11/774,073. Because of vibration and relative motion betweenthe camera and the separation chamber, it is unlikely that the samepixels in the camera will image the observation regions from onerotation to the next. Nevertheless, by selecting pixels with referenceto a dynamically determined origin, a stable view of the observationregions can be obtained.

In the present invention, a series of points representing an edge ismeasured by collecting raw intensity data from pixels in the cameraimaging a region that crosses the edge, filtering the intensity data toreduce variation, and differences between adjacent pixels. There is anabrupt change from light to dark at the edge, which is detected by adifference minima. A set of data points, preferably about five (5), iscollected for each edge, and a line is computed through the points. Anerror measurement, for example, the root mean square error, iscalculated for each line. If the error is too large, the image (or“frame”) for the current rotation is abandoned. The data is deemed tooimprecise or noisy. A new frame would be available in about 40microseconds, and the process can begin again. The line with the leasterror is selected as a referent line. A new or dependant line iscalculated for the line with the greater error. The dependant line is amathematical construct created at the known angle between the two edges.This angle may be any acute or obtuse angle less than 180 degrees. Thepreferred angle is 90 degrees.

The intersection of the two edges is usually physically chamfered. Toprovide a precise intersection of the referent and dependant lines, theyare preferably calculated at an offset from their constituent datapoints. The intersection of the calculated lines will not fall in thechamfer area and data from the chamfered area will be excluded fromcalculations. The error function is again computed for the dependantline. If the error exceeds a selected maximum, the frame is discarded,as described above.

Using the parameters of the lines a transformation function is produced,which translates data points from an (r, s) co-ordinate domain derivedfrom measurements of the edges into an (x, y) co-ordinate domain used toidentify pixels in the observation areas. To test the transformation,the data points for the two edges are translated from the (r, s) domaininto the (x, y) domain and the error function is computed once again. Ifthe error exceeds a maximum error limit, the frame is abandoned.

If the data passes the tests, the pixels falling within the observationregions identified with reference to the origin that has been identifiedas the intersection of the referent and dependant lines are useddetermine the position of phase boundaries and out flow characteristics.The process outlined above and described more completely hereafterallows for a frame by frame determination of the location of an originin the pixel field of the camera and for a determination that the imageis sufficiently clear for the collection of data. Vibration and relativemotion between the rotor and separation chamber and the camera causesthe image detected by the camera to move in the (x, y) plane and to comein and out of focus. The method described allows the apparatus todiscard a frame that is too blurry to provide accurate data and tolocate a consistent origin from frame to frame.

As shown conceptually in FIG. 7, the (r, s) co-ordinate domain is anorthogonal planar reference system in two co-ordinates. Herein, the (r,s) domain is associated with the machined, right-angle reference block214 shown in FIG. 5. The (x, y) co-ordinate domain is also an orthogonalplanar reference system in two co-ordinates. The (x, y) is associatedwith the phase boundary monitoring region 202 or the collect portmonitoring region 204 or both. The origin of the (x, y) domain isusually offset from the origin of the (r, s) domain and the two axis ofthe two domains are not necessarily parallel to each other. The presentinvention develops a transformation of information from one domain tothe other. The transformation takes the following form, which specifiesthe conversion between S, R (pixel) coordinates and X, Y (engineeringunits) coordinates. The matrix is defined as:

$\begin{pmatrix}X \\Y \\1\end{pmatrix} = {T\begin{pmatrix}S \\R \\1\end{pmatrix}}$ $\begin{pmatrix}S \\R \\1\end{pmatrix} = {T^{- 1}\begin{pmatrix}X \\Y \\1\end{pmatrix}}$ $T = \begin{pmatrix}{P_{S}\cos\; a} & {P_{R}\sin\; a} & {{{- P_{S}}D_{S}\cos\; a} - {P_{R}D_{R}\;\sin\; a}} \\{{- P_{S}}\sin\; a} & {P_{R}\cos\; a} & {{P_{S}D_{S}\sin\; a} - {P_{R}D_{R}\cos\; a}} \\0 & 0 & 1\end{pmatrix}$ $T^{- 1} = \begin{pmatrix}\frac{\cos\; a}{P_{S}} & {- \frac{\sin\; a}{P_{S}}} & {- D_{S}} \\\frac{\sin\; a}{P_{R}} & \frac{\cos\; a}{P_{R}} & D_{R} \\0 & 0 & 1\end{pmatrix}$Where P_(S) is the pixel size (in microns) along the S axis, P_(R) isthe pixel size (in microns) along the R axis, a is the angular rotationbetween the S,R and X,Y coordinate systems, D_(S) is the S position ofthe reference corner, and D_(R) is the R position of the referencecorner.

The optical reference 214 has a horizontal edge 232 and a vertical edge234. Preferably, these edges 232, 234 intersect at a right angle,although the angle might also be acute or obtuse. The detectionalgorithm begins 236 by identifying an r Axis Line representative of thehorizontal edge 232. An index i is initialized 238. Data pointsrepresenting detected positions along the edge are added 240 and theindex is incremented 242 until a selected number of data points arecollected 244. Data points represent transitions from dark to lightalong the horizontal edge, as determined by the camera. The process ofselecting data points is represented graphically in FIG. 8. Rawintensity data 252 acquired from pixels in the field of view of thecamera along a line generally parallel to the s axis (that is, radially)for detecting points on the horizontal edge 232 may be filtered 254 toreduce noise related variations. Taking the difference 256 betweenadjacent pixels, moving from light 258 to dark 260, a minimum (absolutevalue maximum) difference 262 is found at the pixels closest to the edge232. A horizontal sweep would be used to locate points on the verticaledge 234. If a satisfactory set of data points is collected, arepresentative line is fitted 264 through the data points. If the datapoints are sufficiently close to the line, for example if the meansquare fit of the data to the line is less than a predetermined limit,the r Axis Line fit criteria is satisfied 266, and the program cancontinue. Otherwise, the program is interrupted 268. An interruption inthe program implies that the apparatus will attempt to acquire asatisfactory image on the subsequent rotation of the centrifuge.

The program then calculates selected characteristics of the r line. Theslope of the line is computed 270. An offset 272 is added to the data.This essentially moves the line representing the horizontal edge 232upward so that the line does not intersect a chamfer at the junction ofthe two edges 232, 234. Data points for the vertical edge will beselected in a region above the offset line. Finally, an errorco-efficient for the data with respect to the r Axis line is computed.This co-efficient may be, for example, the root mean squared error ofthe data with respect to the line.

The detection algorithm next identifies an s Axis Line representative ofthe vertical edge 234. An index i is initialized 276. Data pointsrepresenting detected positions along the edge are added 278 and theindex is incremented 280 until a selected number of data points arecollected 282. Data points represent transitions from dark to lightalong the vertical edge, as described above. If a satisfactory set ofdata points is collected, a representative line is fitted 284 throughthe data points. If the data points are sufficiently close to the line,for example if the mean square fit of the data to the line is less thana predetermined limit, the s Axis Line fit criteria is satisfied 286,and the program can continue. Otherwise, the program is interrupted 288.

The program then calculates the selected characteristics of the s Axisline. The slope of the line is computed 290. An offset 292 is added tothe data. As before, this moves the line representing the vertical edge234 sideways so that the line does not intersect the chamfer at thejunction of the two edges 232, 234. Finally, an error co-efficient forthe data with respect to the s Axis line is computed. This co-efficientmay be, for example, the root mean squared error of the data withrespect to the s Axis line.

The next major feature of the program selects the line having the mostconsistent data, that is data that is most linear, and thenre-calculates the other line at a right angle (or other angle dependingon the optical reference 214) to the selected line. An offset for thecalculated line is selected to minimize error of the data points withrespect to the calculated line. The error is computed, and, if the errorexceeds a pre-determined limit, the program is interrupted in favor ofthe next frame, as explained above. This feature comprises steps 296through 360.

In step 296, the program compares the error for the s Axis line to theerror for the r Axis line. If the error for the s Axis line is smallest,a line will be calculated for the horizontal or r Axis, representing thehorizontal edge 232. The program will calculate an offset for the pointwhere the calculated r Axis will cross the s Axis according to thefollowing formula:

$a = \frac{{\frac{2}{N}{\sum s_{i}}} - {\left( {\frac{2}{N}{\sum r_{i}}}\; \right)m}}{2}$

where a is the offset, N is the number of data points in the selectedline, s_(i) and r_(i) are data points along the vertical and horizontaledges 234, 232 respectively, and m is the slope of the horizontal or rAxis line, set, in this example, to −90 degrees from the s Axis line. Tocalculate the offset a, an index i is set 298 to zero. Position sums(Σs_(i) and Σr_(i)) for the s Axis 300 and for the r Axis 302 areaccumulated for the number of data point along the s Axis and the indexi is incremented 304 until the number of points along the s Axis hasbeen reached 306. The respective position sums are multiplied by 2 anddivided by the number of data of data points along the r Axis 308 andthe s Axis 310, respectively. The r Slope is set 312 to −90 degrees fromthe s Axis line. This slope is forced to match the known included anglein the optical reference 214, in this case, 90 degrees. Of course, otherangles could be used. The offset for the calculated r or horizontal lineis calculated 314 as half of the accumulated s Axis values minus the sumof the r Axis values times the r Slope. The sum for the r Axis and anindex are cleared 316, 318. The error each data point along the r Axis,that is, the deviation of the point from the calculated and offset rLine, is calculated 320 and the square of each deviation is accumulated322. The index is incremented 324 until all points have been accumulated326. The average error for the r Axis points is calculated 328.

On the other hand, if the error for the r Axis line is smallest, a linewill be calculated for the vertical or s Axis, representing the verticaledge 234. The program will calculate an offset for the point where thecalculated s Axis will cross the r Axis. To calculate the offset, anindex i is again set 330 to zero. Position sums (Σs_(i) and Σr_(i)) forthe s Axis 332 and for the r Axis 334 are accumulated for the number ofdata point along the r Axis and the index i is incremented 336 until thenumber of points along the r Axis has been reached 338. The respectiveposition sums are multiplied by 2 and divided by the number of data ofdata points along the r Axis 340 and the Axis 342, respectively. The sSlope is set 344 to −90 degrees from the r Axis line. The offset for thecalculated s or vertical line is calculated 346 as half of theaccumulated r Axis values minus the sum of the s Axis values times the sSlope. The sum for the s Axis and an index are cleared 348, 350. Theerror each data point along the s Axis, that is, the deviation of thepoint from the calculated and offset s Line, is calculated 352 and thesquare of each deviation is accumulated 354. The index is incremented356 until all points have been accumulated 358. The average error forthe s Axis points is calculated 360.

Depending on which Axis line was calculated, the average error foreither the r Axis or the s Axis is compared 362 to a pre-selectedmaximum. If successful, the calculated line will be at right angles tothe other line and will pass through the data points in such a way thatthe error is comparatively low. If the error is too large, however, theprogram interrupts 364 processing and allows the device to try tocapture another image on the next rotation of the centrifuge.

If the lines representing the edges 232, 234 have been successfullyestablished, the program is prepared to call a “get Transform”subroutine 366 to relate the (r, s) co-ordinates to the (x, y)coordinates of the observation regions. An acceptable average error andan acceptable maximum error are selected 368. Parameters P_(s), P_(r),D_(s), D_(r), and the offset “a” are selected 370. The transformpreferably takes the form set forth above.

The program then implements the transform against the data collected forthe horizontal and vertical edges 232, 234 of the marker 214. An errormeasurement is generated, (preferably a root-mean square measurement),to check the efficacy of the transformation. First, for the horizontaledge, an index p is set 372 to zero. For each data point along the xaxis, the data point is transformed 374 into the (x, y) co-ordinatesystem. The error in the data is computed and summed together 376. Ifthe error is greater 378 than the preselected limit, the program goes380 to “BREAK”, and once again waits for the next frame to appear. TheP-counter is incremented 382 until P equals or exceeds 384 the number ofdata points on the x axis.

Next, for the vertical edge, the index p is set 386 to zero. For eachdata point along the y axis, the data point is transformed 388 into the(x, y) co-ordinate system. The error in the data is computed and summedtogether 390. If the error is greater 392 than the preselected limit,the program goes 394 to “BREAK”, and once again waits for the next frameto appear. The P-counter is incremented 396 until P equals or exceeds398 the number of data points on the y axis.

If the signal processing algorithm described above has successfullylocated an origin, that is, the intersection of the edges 232, 234, andthe data has successfully each of the error tests described, and thealgorithm has produced a robust transformation for the frame, theapparatus may begin processing 398 the data in observation regions. Apreferred apparatus and method for such signal processing has beendescribed in U.S. patent application Ser. No. 11/772,692 and U.S. patentapplication Ser. No. 11/774,073, the disclosure of which is incorporatedherein. Such processing would allow the apparatus to distinguish phaseboundaries in the observation region 202 and to distinguish cell typesin the outflow region 204. In response to observed changes in theregions 202, 204, various operating parameters, such as the speed ofperistaltic pumps, may be adjusted to control the operatingcharacteristics of the blood processing apparatus.

The optical reference control described herein allows for frame by framerecognition of the location of important control features within avibrating two-dimensional optical field. Moreover, throughout theprocess of recognizing the reference point, the quality of the visualimage being detected is checked. If the error in the quality of theimage exceeds certain limits (for example, when the data points are notsufficiently linear), the frame is abandoned without further processingin favor of the next frame or visual image. Finally, the transformationitself is checked against data associated with optical reference 214before any attempt is made to process data derived from the observationregions. This assures consistent recognition of the same physicalobservation regions, despite imaging of those regions on differentpixels in the camera. Therefore, sensitive optical recognitiontechniques can be used with respect to the observation regions, in spiteof the vibrations and other optical noise associated with a high speedcentrifuge.

It will be apparent to those skilled in the art that variousmodifications and variations can be made to the structure andmethodology of the present invention without departing from the scope orspirit of the invention. Rather, the invention is intended to covermodifications and variations provided they come within the scope of thefollowing claims and their equivalents.

1. A centrifuge blood processing system for separating fluid componentscomprising: a separation chamber rotating about a central rotation axis,said separation chamber having an optical reference mounted thereon, afirst detector in optical communication with said separation chamber;means for detecting a plurality of observations from an observationregion on said separation chamber with said first detector; acomputational apparatus comprising means for distinguishing said opticalreference and establishing a two-dimensional co-ordinate domain withrespect to said optical reference, means for gathering data correlatedto said co-ordinate domain from said observations of said observationregion as detected by said first detector, and means for controllingfluid flow in said blood processing system in response to said data. 2.The centrifuge blood processing system of claim 1 wherein saidcomputational apparatus comprises means for rejecting an observation ofsaid separation chamber if an error measurement of said opticalreference is not within a pre-selected error limit for said opticalreference.
 3. The centrifuge blood processing system of claim 2 whereinthe computational apparatus comprises means for translating said datafrom said observation region from a first co-ordinate domain into asecond co-ordinate domain thereby producing translated data, and meansfor controlling fluid flow in said blood processing system in responseto said translated data.
 4. The centrifuge blood processing system ofclaim 3 wherein said computational apparatus rejects an observation ofsaid separation chamber if translated data representing said opticalreference is not within a pre-selected error limit for said translateddata.
 5. The centrifuge blood processing system of claim 1 wherein saidoptical reference comprises at least two non-parallel sides and whereinsaid computational apparatus recognizes a first side represented by afirst line and further fits for fitting a second line to a second sideaccording to a known angle between said first side and said second side.6. The centrifuge blood processing system of claim 5 wherein thecomputational apparatus comprises means for translating said data fromsaid observation region from a first co-ordinate domain into a secondco-ordinate domain, thereby producing translated data, and means forcontrolling fluid flow in said blood processing system in response tosaid translated data.
 7. The centrifuge blood processing system of claim6 wherein said computational apparatus comprises means for rejecting anobservation of said separation chamber if translated data representingsaid optical reference is not within a pre-selected error limit for saidtranslated data.
 8. The centrifuge blood processing system of claim 5wherein the computational apparatus comprises means for rejecting anobservation of said separation chamber if data representing an edge ofsaid optical reference are not within a pre-selected error limit forsaid translated data.
 9. The centrifuge blood processing system of claim8 wherein the computational apparatus comprises means for computing anerror measurement for each of said first and second sides and forselecting the side with the least error measurement as a referent line.10. The centrifuge blood processing system of claim 9 wherein thecomputational apparatus comprises means for computing a dependant linefor the side with the greater error measurement.
 11. The centrifugeblood processing system of claim 10 wherein the computational apparatuscomprises means for computing an error measurement for the dependantline and for rejecting an observation of said separation chamber if theerror measurement for the dependant line is not within a pre-selectederror limit for the dependant line.
 12. The centrifuge blood processingsystem of claim 5 wherein the first and second sides are orthogonal toeach other.
 13. The centrifuge blood processing system of claim 12wherein the computational apparatus comprises means for rejecting anobservation of said separation chamber if data representing an edge ofsaid optical reference are not within a pre-selected error limit forsaid translated data.
 14. The centrifuge blood processing system ofclaim 13 wherein the computational apparatus comprises means forcomputing an error measurement for each of said first and second sidesand for selecting the side with the least error measurement as areferent line.
 15. The centrifuge blood processing system of claim 14wherein the computational apparatus comprises means for computing adependant line for the side with the greater error measurement.
 16. Thecentrifuge blood processing system of claim 15 wherein the computationalapparatus comprises means for computing an error measurement for thedependant line and for rejecting an observation of said separationchamber if the error measurement for the dependant line is not within apre-selected error limit for the dependant line.
 17. The centrifugeblood processing system of claim 16 wherein the computational apparatuscomprises means for translating said data from said observation regionfrom a first co-ordinate domain into a second co-ordinate domain,thereby producing translated data.
 18. The centrifuge blood processingsystem of claim 17 wherein said computational apparatus comprises meansfor rejecting an observation of said separation chamber if translateddata representing said optical reference is not within a pre-selectederror limit for said translated data.